The incidence of upper gastrointestinal adenocarcinomas (UGCs;adenocarcinomas of the esophagus and stomach) has been rising steadily. Moreover, a sharp increase in the incidence of lower esophageal and gastroesophageal junction adenocarcinomas has been observed over the past three decades bearing the distinction of the fastest rising incidence of all cancers in the Western world. Overall relative 5-year survival rates are currently less than 20%. Improvement in our presently limited diagnostic, preventive, and therapeutic approach to UGCs is currently a pressing issue. This proposal is based on our original findings through the use of multiple molecular analyses in UGCs that led to the cloning and identification of t-DARPP as a novel cancer gene. We found over-expression of t-DARPP in approximately two-thirds of UGCs and analysis of its biological functions indicated that t-DARPP is a potent pro-survival protein. Using several drugs (camptothecin, butyrates, ceramides) we found that t-DARPP protects cancer cells against drug-induced cell death by a mechanism that includes activation of AKT survival pathway and up-regulation of BCL-2. We hypothesize that t-DARPP provides critical potent pro-survival and anti-apoptotic advantages to cancer cells leading to resistance to drug- induced cell death. We plan to investigate the molecular and biological roles of t-DARPP and determine the potential therapeutic value of its knockdown in UGCs. In Aim 1 of this proposal, we will investigate the molecular mechanism(s) by which t-DARPP phosphorylation sites regulate the AKT survival pathway. Based on our preliminary data, we will investigate the mechanisms by which t-DARPP regulates the PTEN tumor suppressor activity, localization, and stability in cancer cells. In Aim 2, we plan to investigate the effect of t- DARPP on drug resistance by testing its ability to abrogate drug-induced cell death. We will examine the role of t-DARPP in regulating the complex intrinsic apoptosis. Several recent reports suggest the oncogene addiction concept in cancer cells, where cancer cell survival becomes dependent on the expression of a critical survival protein. We will test the therapeutic potential of t-DARPP knockdown in vivo and whether its knockdown alone or in combination with the existing chemotherapeutics can boost the therapeutic response. In Aim 3, we will adopt powerful proteomic approaches for a systematic characterization of proteins that interact with t-DARPP and those that are downstream effectors in order to identify novel signaling pathways and biological functions that t-DARPP could mediate in cancer cells that remain unknown. In all our experiments, we will test its phosphorylation mutants in order to characterize the molecular and functional role(s) of each of these phosphorylation sites in cancer cells. We anticipate that completion of the suggested experiments will provide a significant insight into the role of t-DARPP, as a novel protein, in upper gastrointestinal adenocarcinomas. PUBLIC HEALTH RELEVANCE: This proposal connects in vitro and in vivo experiments in order to characterize the role(s) of t-DARPP in upper gastrointestinal carcinogenesis. We plan to investigate the role of t-DARPP in regulating the intrinsic apoptosis and evaluate its potential as a novel therapeutic target. State-of-the-art proteomic approaches will be employed to identify t-DARPP interacting proteins as well as its molecular signaling targets.